Screening Technique and its Details
The screening techniques are generally carried out in two stages.
- Primary Screening
- Secondary Screening
1. Primary Screening
Detection and isolation of industrially important micro-organism from the mixed population using simple techniques is called as primary screening techniques.In primary screening technique, simple methods are used to detect valuable micro-organism based on certain biochemical characters of microbes.Generally, the sample is collected, serially diluted and isolated on suitable agar medium by a specific technique.
Here are some examples of primary screening
a] Screening of antibiotic-producing micro-organism.
In the screening of antibiotic-producing micro-organism, a crowded plate technique is used.Here source used for screening of micro-organism is generally soil.The sample of soil used is serially diluted in sterile distil water.The suitable amount of diluted sample that is 0.1 ml is taken by a sterile pipette and spread or sprayed on suitable sterile agar medium in sterile conditions.This inoculated medium is incubated in incubators at room temperature for 24 to 48 hours.These cultures are in incubated at room temperature because the source of micro-organism is soil and room temperature is suitable for growth of this microbes.The agar plates which have 300 to 400 colonies are selected.The plates having well-isolated colonies are used for further studies as the well-isolated colonies show the clear zone due to the production of antibiotic and inhibition of growth of another micro-organism.These antibiotic-producing colonies are further sub-cultured on fresh desired media and sub-cultured growth is purified by re-streaking.The stock cultures are maintained and used for further studies.
b] Screening of organic acid or amine producing micro-organism
In the screening of organic acid or amine producing micro-organism the source used may be soil, milk or milk product.The source taken is diluted by serial dilution technique in sterile distilled water and spread inoculated on agar medium.These inoculated plates are incubated for suitable incubation period at optimum temperature.After incubation well-isolated colonies are selected.
The nutrient medium used for isolation should contain all required nutrients, a source of carbon and nitrogen.Along with this, the agar medium should contain a pH indicator dye like neutral red or bromothymol blue and the media should be poorly buffered so that the minor changes in pH of media and production of amine or acid can be detected. After incubation of Inoculated plates, the production of acid or amine is detected by the change in colour of media surrounding the colony.The change of colour of media depends on the production of acid or amine.
There is one more method that can be used for screening of organic acid producing micro-organism and that is the addition of calcium carbonate in the nutrient media instead of pH indicator dye.The initial method of inoculation is same as above after inoculation the organic acid producing colony shows a clear zone around colony as the organic acid produced by colony dissolves the calcium carbonate present in media.Thus the organic acid producing colonies are further sub-cultured, purified and stock culture is maintained.
c] Screening of organism producing Vitamin, Amino acids and other Growth factors.
Here for a screening of organisms, a suitable source is selected and serial dilution is made in sterile distilled water.This dilution is inoculated on a suitable nutrient agar medium without any vitamin, amino-acid or metabolite. The nutrient media should contain all required essential nutrients except the metabolite or growth factor that is under consideration.The inoculated plates are incubated for a suitable period of time at optimum temperature.After incubation, the plates are taken and the colonies obtained may be vitamin, amino-acid or metabolite producer.
The isolated colonies are check for production of metabolites whether the metabolites are produced in excess because the organism producing these metabolites in excess utilize some quantity of metabolites for its growth and an excess amount of metabolites are secreted extracellularly in the vicinity of the colony.
In primary screening, we detect and isolate the desired organism but in secondary screening, we characterize industrially important micro-organism which is isolated in primary screening by using highly selective procedures. Here in secondary screening, we screen the organism that is capable of giving the high yield of the product by using the cheap raw material.
In primary screening, only basic information about the isolated colony is obtained and in secondary screening, the detail information about the isolate is obtained and it is determined whether the isolate is capable of being used on industrial scale. Before using the isolate on industrial scale determination of its capabilities and yield potential is important so secondary screening plays a very important role. The valueless organism is discarded only valuable organism are used further as secondary screening may involve expensive procedure of screening. The organism suitable for commercial production of the product is detected and used further.
Secondary screening is generally used to obtain information about’isolated micro-organism.The information obtained is as follows:-
- It can use to determine qualitative as well as quantitative information about strains.Qualitative in the sense determination of inhibitory spectrum in case of antibiotics and yield potential of the product and quantitative is the determination of the quantity of product obtained by using different fermentation media.
- Secondary screening helps to determine the product produced by fermentation media by using various techniques like chromatography.
- It provides information necessary for classification and identification of organism and due to which the pathogenicity to human and animals can be determined.
- Secondary screening helps to determine the genetic instability of strain.It is very important because if organism carries out mutation and loses the capabilities of giving high yield then the industry may face a great loss.
- In secondary screening, various nutrients that are provided for growth are tested for toxicity so that any nutrient compound that is toxic to growth of isolated strain should be eliminated.
- The chemical solubility of product produce is tested.The organic solvents in which the product can get dissolved are eliminated.
- The physical, chemical and biologically properties of the product are determined and studied.
- The secondary screening determines the capabilities of an organism to alter or destroy its own product.
Thus secondary screening is all about the collection of a broad range of information about the isolated strain and on the bases of this information, it is decided whether the selected micro-organism is suitable for use on an industrial scale or not.